C7790 Introduction to Molecular Modelling -1Lesson 2 Computational Chemistry vs Experiment Petr Kulhánek kulhanek@chemi.muni.cz National Centre for Biomolecular Research, Faculty of Science Masaryk University, Kamenice 5, CZ-62500 Brno JS/2022 Present Form of Teaching: Rev5 C7790 Introduction to Molecular Modelling TSM Modelling Molecular Structures C9087 Computational Chemistry for Structural Biology C7790 Introduction to Molecular Modelling -2Method overview (model chemistry] Quantum mechanics Molecular mechanics Coarse-grained mechanics atomic resolution bead resolution reactivity domain movement, folding atomic resolution bead resolutionatomic resolution bead resolution conformational movements up to 1,000 atoms * up to 1,000,000 beads *up to 1,000,000 atoms * up to 100 ps * up to ms *to 1 ms * C7790 Introduction to Molecular Modelling -3Importance of computational chemistry computational chemistry experiment theory "resolution”"accuracy” Computational chemistry can work with single atom resolution. C7790 Introduction to Molecular Modelling -4Atomic resolution computational chemistry experiment atomic resolution since the introduction of quantum theory (1925) ➢ it refines models ➢ it improves calculation procedures ➢ it achieves more accurate results in less computational time atomic resolution since the introduction of X-ray crystallography (1923) ➢ it refines techniques ➢ it improves the resolution Experiments with single atom or molecule resolution. (Single Molecule Experiments) Historicaldevelopment C7790 Introduction to Molecular Modelling -5Atomic Resolution Experiments C7790 Introduction to Molecular Modelling -6X-ray Crystallography X-ray striking an electron produces secondary spherical waves emanating from the electron. This phenomenon is known as elastic scattering, and the electron is known as the scatterer. A regular array of scatterers produces a regular array of spherical waves. Although these waves cancel one another out in most directions through destructive interference, they add constructively in a few specific directions, determined by Bragg's law: Diffraction pattern (enzyme crystal) http://www.wikipedia.org  nd =sin2 Disadvantages: • the sample must be a monocrystal • radiation damage X-rays diffracts on electrons from atoms. Diffraction (schematic model) C7790 Introduction to Molecular Modelling -7X-ray Crystallography X-ray crystallography method determines the position of individual atoms in the unit cell of crystal. However, the positions of some atoms may not be determined in the case of low resolution or internal disorder. This usually happens for hydrogen atoms (weakly diffracting), side chains in biomolecules, or weakly bound substrates. Diffraction on crystalls can be achieved with other sources of beams with suitable wavelengths: • Neutrons - Benefit of neutron diffraction is that the diffraction occurs at the nuclei of individual atoms. This method can determine hydrogen atom positions, because protons (hydrogen atom nuclei) difracts very well. • Electrons - electron crystallography, availbale in modern electron microscopes C7790 Introduction to Molecular Modelling -8Nuclear Magnetic Resonance - NMR ➢ chemical shift ➢ J-coupling ➢ NOE (Nuclear Overhauser Effect) - proportional to the distance ➢ and more Advantages: • sample in solution • non-destructive Disadvantages: • isotope labeling • not suitable for very large molecules C7790 Introduction to Molecular Modelling -9Nuclear Magnetic Resonance - NMR NMR spectra Macek, P .; Hops, J .; Cross, I .; Savoy cabbage, P .; Padrta, P .; Žídek, L .; Wild, M .; Hadravová, R .; Chaloupková, R .; Pichová, I .; et al. NMR Structure of the N-Terminal Domain ofCapsid Protein from the Mason – Pfizer Monkey Virus. Journal of Molecular Biology 2009, 392, 100–114. molecularly dynamic simulationexperimental data providing some interatomic distances the resulting structure is represented by several conformations the structure contains hydrogen atoms, which are provided by used theoretical model (molecular mechanics) stretched structure C7790 Introduction to Molecular Modelling -10Scanning Tunneling Microscopy STM http://www.wikipedia.org Principle: Result: Disadvantages: • electroconductive materials C7790 Introduction to Molecular Modelling -11Single Molecule Experiments C7790 Introduction to Molecular Modelling -12Atomic Force Microscopy - AFM Principle: Result: http://www.wikipedia.org C7790 Introduction to Molecular Modelling -13FRET Experiments FRET: Fluorescent Resonance Energy Transfer two chromophores we can determine the distance Principle: Result: BsoBI Q: Is it opened during DNA binding and if so, on which side? C7790 Introduction to Molecular Modelling -14Magnetic and Optical Tweezers Principle: http://www.wikipedia.org Suitable for: ➢ Active/Binding site location ➢ Kinetics measurements C7790 Introduction to Molecular Modelling -15Optical tweezers - use VU University, Amsterdam C7790 Introduction to Molecular Modelling -16Electron cryomicroscopy - cryoEM Building E35/ CEITEC Acceleration voltage: 300 kV Electron microscopy is a form of transmission electron microscopy where a sample is studied at low temperatures (typically liquid nitrogen temperature). The technique is used in structural biology. C7790 Introduction to Molecular Modelling -17Electron cryomicroscopy - cryoEM http://proj.ncku.edu.tw/research/commentary/e/20080919/2.html C7790 Introduction to Molecular Modelling -18Small-angle X-ray scattering SAXS https://wiki.anton-paar.com Small-angle X-ray scattering (SAXS) is technique by which nanoscale density differences in a sample can be quantified. It can determine nanoparticle size distributions, resolve the size and shape of (monodisperse) macromolecules, determine pore sizes, characteristic distances of partially ordered materials, and much more. C7790 Introduction to Molecular Modelling -19Structure Databases It contains about half a million structures of small molecules determined by Xray and neutron diffraction. Suitable software: Mercury http: // www.ccdc.cam.ac.uk/Solutions/CSDSystem/Pages/Mercury.aspx Cambridge Structural Database (CSD) http://www.ccdc.cam.ac.uk/Solutions/CSDSystem/Pages/CSD.aspx It contains about 94 thousand structures of biomolecular systems determined mainly by X-ray structural analysis. Protein Data Bank (PDB) http: //www.pdb.org Experimental method Proteins (P) Nucleic acids (NA) P / NA complexes Other Overall X ray 77445 1481 4069 3 82998 NMR 8851 1046 193 7 10097 electron microscopy 469 45 129 0 643 status in September 2013 C7790 Introduction to Molecular Modelling -20- Summary ➢Use molecular modelling for problems that cannot be solved by experimental techniques ➢Use molecular modelling to complement experimental data ➢NMR, FRET, cryoEM, SAXS, etc. C7790 Introduction to Molecular Modelling -21- Homework C7790 Introduction to Molecular Modelling -22- Homework 1. What is the typical wavelength of radiation used in X-ray structural analysis? 2. What is the de Broglie wavelength of electrons in electron microscopy for an accelerating voltage of 300 kV? 3. How is the fluorescently labeled enzyme BsoBI prepared (page 13)? 4. How many structures determined by electron microscopy are currently stored in the PDB database?